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Biosynthesis of 2‐hydroxyacid‐containing polyhydroxyalkanoates by employing butyryl‐CoA transferases in metabolically engineered Escherichia coli

04:27 EDT 2 Sep 2017 | Wiley Biotechnology Journal

We previously reported the production of polyhydroxyalkanoates (PHAs) containing 2‐hydroxyacid monomers by expressing evolved Pseudomonas sp. 6‐19 PHA synthase and Clostridium propionicum propionyl‐CoA transferase in engineered microorganisms. Here, we examined four butyryl‐CoA transferases from Roseburia sp., Eubacterium hallii, Faecalibacterium prausnitzii, and Anaerostipes caccae as potential CoA‐transferases to support synthesis of polymers having 2HA monomer. In vitro activity analyses of the four butyryl‐CoA transferases suggested that each butyryl‐CoA transferase has different activities towards 2‐hydroxybutyrate (2HB), 3‐hydroxybutyrate (3HB), and lactate (LA). When Escherichia coli XL1‐Blue expressing Pseudomonas sp. 6‐19 PhaC1437 along with one butyryl‐CoA transferase was cultured in chemically defined MR medium containing 20 g/L of glucose, 2 g/L of sodium 3‐hydroxybutyrate, and various concentrations of sodium 2‐hydroxybutyrate, PHAs consisting of 3HB, 2HB, and LA were produced. The monomer composition of PHAs agreed well with the substrate specificities of butyryl‐CoA transferases from E. hallii, F. prausnitzii, and A. caccae, but not Roseburia sp. When E. coli XL1‐Blue expressing PhaC1437 and E. hallii butyryl‐CoA transferase was cultured in MR medium containing 20 g/L of glucose and 2 g/L of sodium 2‐hydroxybutyrate, P(65.7 mol% 2HB‐co‐34.3 mol% LA) was produced with the highest PHA content of 30 wt%. Butyryl‐CoA transferases also supported the production of P(3HB‐co‐2HB‐co‐LA) from glucose as the sole carbon source in E. coli XL1‐Blue strains when one of these bct genes was expressed with phaC1437, cimA3.7, leuBCD, panE, and phaAB genes. Butyryl‐CoA transferases characterized in this study can be used for engineering of microorganisms that produce PHAs containing novel 2‐hydroxyacid monomers.

Original Article: Biosynthesis of 2‐hydroxyacid‐containing polyhydroxyalkanoates by employing butyryl‐CoA transferases in metabolically engineered Escherichia coli

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